Many patients with severe dermal burn injury develop an exaggerated healing response resulting in hypertrophic scarring. This excess scar results in limited mobility, joint contracture and gross disfigurement. Treatment modalities are limited. Hence, hypertrophic scar is the major factor limiting complete recovery from burn injury. Previous investigations and preliminary data suggest that hypertrophic scar tissue and fibroblasts derived there from exhibit higher rates of collagen synthesis and collagen lattice contraction relative to unscarred skin. Fibroblasts grown from hypertrophic scars have slower growth rates and are less responsive than normal skin cells to serum and EGF. In addition, preliminary experiments have shown a disordered interaction between keratinocytes and fibroblasts and excess early cytokine release, in particular TGFbeta. Hence, our working hypothesis is that alterations in early burn healing induce both increased extracellular matrix production and abnormal cytokine production. These result in stable phenotypic alterations within the wound which are manifested by excess collagen synthesis, as well as abnormal autocrine and paracrine responses to selected cytokines. The net overall result is to promote exuberant scar formation. The experimental strategy is based on three primary aims. First, cytokine abnormalities of the burn would will be characterized using biochemical and molecular biologic approaches. In particular, the role of TGFbeta as the proximate cause of increased matrix production will be documented. Second, the phenotypic alterations in hypertrophic scar tissue (ex vivo) and cells (keratinocytes and fibroblasts (in vitro) will be characterized in terms of collagen and cytokine (TGFbeta) gene expression using biochemical, molecular biological and immunological approaches. The response of scar fibroblasts and keratinocytes to exogenous pure cytokines (EGF, PDGF, IL-8, TNFalpha, TGFbeta) will be compared to both those derived from injured tissues which heal without excess scar and normal cells from the same individual. Finally, the directional regulatory influences keratinocyte-derived cytokines on fibroblasts will be examined in vitro using conditioned medium, and the responsible agents identified using chromatographic and immunological approaches. The functional end points to be studied include cellular growth rate, collagen production, and the production of cytokines. In this matter the phenotype alterations in hypertrophic scar will be delineated and the role of cytokines in their causation determined. The long-term goal of this project is to uncover key mechanisms underlying hypertrophic scarring which may lead to improved therapeutic and perhaps preventive strategies.